J Periodontol August 2008 (Suppl.

The Role of Inammatory Cytokines in Diabetes and Its Complications

George L. King*
The prevalence of diabetes worldwide is increasing rapidly in association with the increase in obesity. Complications are a major fear of patients with diabetes. Complications of diabetes affect many tissues and organs, causing retinopathy, nephropathy, neuropathy, cardiovascular diseases, peripheral vascular diseases, stroke, and periodontal pathologies. Immunologic abnormalities are associated with type 1 and type 2 diabetes and diabetic complications. T cell abnormalities are believed to be the major cause of autoimmune disease in type 1 diabetes, leading to the destruction of pancreatic islets. In type 2 diabetes, inammation and activation of monocytes are postulated to be important for enhancing insulin resistance and may contribute to the loss of insulin secretory function by islet cells. Many factors can enhance insulin resistance, including genetics, a sedentary lifestyle, obesity, and other conditions, such as chronic inammation or infection. Increases in inammation, such as activation of monocytes and increased levels of inammatory markers, e.g., C-reactive protein, plasminogen activator inhibitor-1, and other cytokines, were reported in insulin-resistant states without diabetes. One possible mechanism is that abnormal levels of metabolites, such as lipids, fatty acids, and various cytokines from the adipose tissue, activate monocytes and increase the secretion of inammatory cytokines, enhancing insulin resistance. According to this model, obesity activates monocytes and enhances insulin resistance, increasing the risk for type 2 diabetes. Abnormalities in innate immunity might also participate in the development of diabetic complications. In general, hyperglycemia is the main initiator of diabetic retinopathy, nephropathy, and neuropathy, and it participates in the development of diabetic cardiovascular diseases. Although the precise role of inammation in the development of diabetic microvascular diseases is still unclear, it is likely that inammation induced by diabetes and insulin resistance can accelerate atherosclerosis in patients with diabetes. Also, it was shown that conditions with an inammatory basis, such as obesity and type 2 diabetes, can contribute to periodontal disease, suggesting that periodontal abnormalities may be partly inuenced by inammatory changes. Further research is required to conrm the role of inammation and the onset of diabetes, microvascular diseases, and periodontal pathologies. J Periodontol 2008;79:1527-1534. KEY WORDS Complications; diabetes; hyperglycemia; inammation; insulin resistance; periodontal disease.

* Section on Vascular Cell Biology, Joslin Diabetes Center, Boston, MA. Department of Medicine, Harvard Medical School, Boston, MA.

iabetes is a growing concern. Its incidence is increasing rapidly and is predicted to increase further, in parallel with the trends observed for obesity.1-3 The increasing prevalence of diabetes represents a signicant burden to human health because of its numerous and often serious complications. These include nephropathy, retinopathy, neuropathy, cardiovascular disease, and periodontitis.4 Diabetes also has an economic cost, with total direct and indirect medical costs already >$132 billion in the United States alone.5 Many factors are known to contribute to the development of diabetes and its complications. These include genetics, diet, sedentary lifestyle, perinatal factors, age, and obesity.6 Nevertheless, an inammatory basis for diabetes and its complications has been gaining interest. Inammatory processes are associated with type 1 and type 2 diabetes; however, the distinct etiology of the two types of diabetes suggests that different causal mechanisms are involved. Type 1 diabetes is frequently found in childhood or young adulthood and arises from the autoimmune destruction of the pancreatic islet cells leading to loss of insulin production.5 Type 2 diabetes is the more common form and occurs mainly in
doi: 10.1902/jop.2008.080246

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adults, although the prevalence in younger people is beginning to increase in conjunction with childhood obesity.5 Type 2 diabetes is characterized by an increase in insulin resistance in conjunction with the inability of pancreatic beta cells to secrete sufcient insulin to compensate.5 Investigations into the role of inammatory mechanisms in diabetes and its complications are expected to provide insight into the processes underlying the onset and progression of the disease. Such improved understanding of the inammatory basis for diabetes may prove valuable for introducing novel approaches to treatment, alongside currently used non-pharmacologic and pharmacologic interventions. INFLAMMATORY PROCESSES ARE ASSOCIATED WITH PROGRESSION OF TYPE 1 DIABETES Although there is some controversy surrounding the precise role of inammatory processes in type 1 diabetes, intriguing ndings have emerged from studies of C-reactive protein (CRP) levels, a measure of circulating inammatory biomarkers.7 Although CRP concentrations in individuals with the new onset (within days of diagnosis) of type 1 diabetes were similar to those observed in healthy controls, levels in individuals with long-term diabetes were signicantly higher (P = 0.04).7 These ndings suggest that the inammatory process may play a greater role in the long-term progression of type 1 diabetes than in its onset. The strongest factor to support inammation as being important in type 1 diabetes is in the area of complications. Increases in inammatory and oxidative stress markers are also found in conjunction with the development of complications of diabetes, with increases in plasma levels of CRP and in concentrations of soluble vascular cell adhesion molecule-1 and nitrotyrosine in patients with microvascular disease compared to those without.8 Increases in monocyte release of interleukin (IL)-1b and superoxide anions were also reported in patients with type 1 diabetes, suggesting the elevations in inammatory marker activity in microvascular and cardiovascular diseases.8 INFLAMMATORY PROCESSES ARE ASSOCIATED WITH DEVELOPMENT OF TYPE 2 DIABETES Research has focused more on the role of inammatory processes in the development and progression of type 2 diabetes than type 1 diabetes. Increases in inammatory markers are detected in apparently healthy individuals who later go on to develop type 2 diabetes,9-11 suggesting that inammation occurs early during the period of impaired glucose tolerance, prior to the diagnosis of type 2 diabetes. In adult Pima Indians, a population characterized by a high prevalence of type 2 diabetes, individuals with white blood cell counts within the highest tertile were more
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likely to develop type 2 diabetes over the 20-year period studied compared to those in the lowest tertile.9 Similarly, in a prospective, nested case-control study10 of apparently healthy, middle-aged women in the United States, inammatory markers IL-6 and CRP within the highest quartiles were associated with an increased risk for developing type 2 diabetes over a 4-year period compared to those in the lowest quartile (unadjusted relative risk [RR], 7.5; [95% condence interval (CI): 3.7 to 15.4] for IL-6 and 15.7 [95% CI: 6.5 to 37.9] for CRP; P <0.001 for both). These ndings were mirrored in the Monitoring of Trends and Determinants in Cardiovascular Diseases study of healthy, middle-aged men, in which CRP concentrations in the highest quartile were associated with an increased risk for developing type 2 diabetes over the 7-year period studied (unadjusted RR, 2.84 [95% CI 1.5 to 5.36]; P = 0.003 for trend).11 INFLAMMATORY PROCESSES CONTRIBUTE TO INSULIN RESISTANCE IN TYPE 2 DIABETES Insulin resistance begins prior to the onset of type 2 diabetes, at which time impaired glucose tolerance occurs as a result of beta cell decomposition and relative insulin deciency. Several factors are linked to the development of insulin resistance in individuals with impaired glucose tolerance and type 2 diabetes, including genetics and environmental inuences, obesity, and other conditions associated with chronic inammation or infection. The possibility that obesity, and the activation of adipose tissue in particular, may enhance the release of inammatory factors that underlie the development of insulin resistance has generated intense interest in the eld of diabetes for a number of reasons. First, a signicant proportion of individuals with type 2 diabetes are overweight or obese, and obesity is a risk factor for the development of type 2 diabetes.2,12 Second, the increased release of adipocyte-derived metabolites, such as lipids, fatty acids, and various inammatory cytokines, in obese individuals has been linked to the development of insulin resistance.13 Third, chronic inammation is associated with obesity, insulin resistance, and type 2 diabetes, all of which are features of the clustering of metabolic pathologies known as metabolic syndrome.14 It should be emphasized, however, that despite interest in obesity as a predisposing factor to insulin resistance in type 2 diabetes, other equally important mechanisms for the loss of insulin sensitivity have been proposed. One study15 of young, lean offspring of patients with type 2 diabetes demonstrated similar body mass index measurements and plasma concentrations of inammatory markers tumor necrosis factor-alpha (TNF-a), IL-6, and adiponectin in insulin-resistant and insulin-sensitive individuals. This suggested that obesity and systemic

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inammatory factors do not play a signicant role in the development of insulin resistance in this population; the loss of insulin sensitivity was ascribed to a dysregulation of intramyocellular fatty acid metabolism. Other studies5,16 demonstrated a high risk for type 2 diabetes in Asian subjects, despite low concentrations of CRP and other inammatory markers compared to other ethnic groups. Taken together, such ndings suggest that mechanisms in addition to obesity and systemic inammation are involved in predisposing individuals to insulin resistance and type 2 diabetes. In the obesity-related model for the development of insulin resistance, adipocytes, once activated, release abnormal levels of bioactive molecules, such as lipids, fatty acids, monocyte chemoattractant protein-1 (MCP-1), and various inammatory cytokines, e.g., CRP, plasminogen activator inhibitor-1, and TNF-a.13 The release of these cytokines and other mediators results in the local recruitment of monocytes within adipose tissues. With differentiation of the monocytes into macrophages comes an increased release of inammatory factors and chemokines locally within adipose tissue and systemically, such that the inammatory response is propagated to various tissues.13 One of the earliest studies17 to link the release of inammatory substances from adipose tissues to insulin resistance in type 2 diabetes involved rodent models of obesity and diabetes. In these mouse and rat models, expression of TNF-a mRNA and protein was induced locally within adipose tissue, as well as systemically in the plasma. By inhibiting TNF-a expression in one of the rodent models (fa/fa) using a recombinant TNF-a receptorimmunoglobulin G chimeric protein, insulin sensitivity improved, suggesting a direct role for TNF-a in the development of insulin resistance.17 Such increases in insulin sensitivity with the use of a specic TNF-a inhibitor have not been replicated in human clinical trials,18 although larger studies and large trial periods may be needed to conrm the ndings.14 A mechanism has been proposed linking the expression of TNF-a and other inammatory mediators to the development of insulin resistance in obesity and type 2 diabetes.13 In this model, inammatory cytokines and/or bacterial lipopolysaccharide stimulate I-kappa-B (IkB) kinase-b (IKKb), and possibly IKKa, to induce activation of nuclear factor-kappa B (NF-kB) (Fig. 1). Other non-inammatory cell/cytokine-mediated agents also contribute to NF-kB activation, including free fatty acids, obesity, hyperglycemia, protein kinase C (PKC) activators, and oxidants. Following activation, NF-kB translocates to the nucleus, resulting in the subsequent transcription of genes that promotes the development of insulin resistance.13 Induction of

target genes by NF-kB leads to the increased expression of inammatory markers and mediators associated with insulin resistance and is associated with the production of other factors, including receptors, inammatory mediators, chemokines, and transcription factors; among other functions, these stimulate recruitment of monocytes and their differentiation into macrophages (Fig. 1).13 INHIBITION OF IKKb/NF-kB PATHWAY IMPROVES OBESITY-RELATED INSULIN SENSITIVITY Several lines of evidence support the critical role of the IKKb/NF-kB pathway in the development of insulin resistance, including a study in an animal model19 and a human clinical trial.20 The heterozygous deletion of IKKb in an obese mouse model fed a high-fat diet over 23 weeks postnatally conferred protection from the development of insulin resistance, as measured by lower fasting insulin and glucose concentrations compared to normal/homozygous IKKb controls.19 Additionally, in a clinical study20 of young, obese, non-diabetic adults, treatment with salsalate, a non-acetylated salicylate and known disruptor of the IKKb/NF-kB pathway, improved insulin sensitivity compared to placebo control. Salsalate treatment also improved inammatory markers in these patients, resulting in a reduction in free fatty acids (P <0.05) and increases in levels of the anti-inammatory cytokine adiponectin (P <0.01) and circulating concentrations of CRP (P <0.05).20 ASSOCIATION BETWEEN INFLAMMATORY PROCESSES AND COMPLICATIONS OF DIABETES Insulin resistance and insulin deciency give rise to a hyperglycemic state that is a major risk factor for the development of diabetic complications. For example, primary pulmonary disorder is rare in diabetes, although the pulmonary tree is heavily vascularized and exposed to all the systematic factors of diabetes. Hyperglycemia is considered the key contributor to microvascular complications, including retinopathy, neuropathy, and nephropathy. It is also one of several major risk factors associated with cardiovascular disease along with insulin resistance or deciency, free fatty acidemia, hypertension, hyperlipidemia, and inammation. Periodontitis is commonly associated with diabetes, as demonstrated by a 2.9-fold increase in risk in individuals with poorly controlled glycemia;21 however, the major risk factors contributing to its development are unknown. Hyperglycemia was shown to act deleteriously through a number of pathways, including the aldose reduction pathway, advanced glycation end product
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Figure 1.
Activators of IKK/NF-kB pathway inhibit insulin signaling. IKK = IkB kinase; IRS = insulin receptor substrate.

contribute to cellular dysfunction and damage associated with micro- and macrovascular complications. The actions of inammatory and oxidant pathways at the local tissue level are key to understanding their contribution to the pathogenesis of diabetic complications.23 Evidence suggests that increases in systemic markers of inammation, such as CRP and IL-6, are associated with complications such as diabetic nephropathy24 in type 2 diabetes. However, systemic inammatory factors are only weakly associated with the development of diabetic retinopathy,25,26 and the relationship is uncertain in other complications, such as periodontal disease. Such studies underscore the importance of investigating local, downstream pathways and mediators for resolving processes underlying vascular pathologies in diabetes.23

LOCAL MECHANISMS FOR DIABETIC COMPLICATIONS Hyperglycemia induces the expression of the PKC pathway.23 This results in the development of complications through altered gene expression and/ or protein function, thus contributing to cellular dysfunction and damage.23 A well-described pathway for the development of diabetic vascular complications involves activation of the diacylglycerol (DAG)-PKC pathway.23,27 Increases in DAG levels and PKC activity are found in a wide variety of tissues and cultured cells isolated from diabetic animals and humans exposed to high glucose levels.27 Several processes may underlie the increased Figure 2. formation of DAG, including the inducMajor pathways initiated by hyperglycemia that contribute to complications of diabetes. tion of oxidants, such as H2O2, and the 22 Reprinted with permission from Blackwell Publishing. accumulation in AGEs (Fig. 3A). Stimulation of DAG synthesis activates PKC (AGE) pathway, reactive oxygen intermediate pathisoforms, with PKC-b considered to play a central role way, and PKC pathway (Fig. 2).22 All four pathways in the development of complications in tissues such as give rise to oxidant and inammatory mediators that retina, kidney, heart, and the vasculature.23,27 Activaresult in deleterious effects, both systemically and lotion of PKC in the vasculature has been associated cally in the tissues. Activation of the pathways leads to with processes including increases in basement enhanced production of gluco-oxidants and AGEs, as matrix protein synthesis, activation of leukocytes, enwell as increased ux through the aldose reduction dothelial cell activation and proliferation, smooth pathway and activation of signaling cascades by memuscle cell contraction, endothelial permeability, acdiators such as PKC, mitogen-activated protein kitivation of cytokines, transforming growth factor-b nase (MAPK), and NF-kB.22 The effects include (TGF-b), vascular endothelial growth factor (VEGF), altered gene expression and/or protein function that endothelin, and angiogenesis.27-29
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Figure 3.
A) PKC-b pathway to diabetic vascular complications. B) Selective local insulin resistance in cardiovascular tissues in combination with the activation of PKC (especially the b isoform) and MAPK inhibits the anti-atherogenic pathway while enhancing proatherogenesis. C and D) Diabetic nephropathy and retinopathy progress through a series of hyperglycemia-dependent pathways in which activation of PKC isoforms enhances activation and/or production of intermediaries, resulting in pathologic changes. The precise role of inammatory cytokines in the progression of retinopathy remains to be determined. Ang = angiotensin; CAD = coronary artery disease; CTGF = connective tissue growth factor; CVD = cardiovascular disease; ET-1 = endothelin-1; eNOS = endothelial nitric oxide synthase; FFA = free fatty acid; Grb-2 = growth factor receptor-bound protein 2; HO-1 = heme-oxygenase-1; IkB = IkB kinase; INS/Ins = Insulin; IRS = insulin receptor substrate; MCT = mean circulation time; PDGF = platelet-derived growth factor; PDK 1 = phosphoinositide-dependent kinase 1; PI3 K P85 = phosphoinositide-3 kinase p85 expression; PKB = protein kinase B; PVD = peripheral vascular disease; RBF = retinal blood ow; Shc = src homology domain c-terminal adaptor homolog.

Vascular complications arise from increased PKC and MAPK activity in cardiovascular tissues in the presence of selective insulin resistance.23,27 Under normal physiologic conditions, insulin interacts with insulin receptors to stimulate two main pathways in cardiovascular tissues: a phosphoinositide-3 kinase (PI3K) pathway that inhibits atherogenesis and has antiatherogenic effects and a MAPK-activated pathway that promotes cellular growth and enhances atherogenesis (Fig. 3B).23,27 In the presence of insulin resistance and diabetes, the increase in glucose and free fatty acids leads to an increased release of inammatory cytokines and altered regulation of PKC and MAPK activity.23,27 PKC inhibits the PI3K pathway and results in enhanced atherogenesis through such processes as a reduction in antiatherogenic nitric oxide production and impaired endothelium-dependent vasodilation. In addition, processes initiated by insulin resistance and diabetes stimulate the MAPK pathway to exert proatherogenic actions. Several lines of evidence suggest that the selective activation of PKC-b is also a likely mechanism for the

pathogenesis of microvascular complications in diabetes, including diabetic retinopathy and nephropathy (Figs. 3C and 3D). In diabetic nephropathy, hyperglycemia is believed to induce damage by activation of the PKC pathway, resulting in proteinuria, mesangium expansion, and nephromegaly followed by glomerularsclerosis. In retinopathy, PKC activation leads to endothelial dysfunction, followed by pericyte loss, formation of acellular capillaries, and microaneurysms. Such pathologic changes in the retina lead to increased vascular leakage and diabetic macular edema (DME) or to increased VEGF and angiogenesis associated with proliferative diabetic retinopathy. Increased VEGF expression is considered a major cause of diabetic proliferative disease30 and may contribute to DME through the stimulation of PKC. SELECTIVE INHIBITION OF PKC-b AMELIORATES THE VASCULAR COMPLICATIONS OF DIABETES With ndings that PKC-b may play a critical role in the development of pathologies associated with insulin
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resistance and diabetes, there is increased interest in the benets of inhibiting PKC-b to reduce harmful sequelae in micro- and macrovascular complications. Studies23,31 have focused on the orally administered selective inhibitor of PKC-b, ruboxistaurin mesylate (RBX; LY333531). RBX is currently in Phase II/III clinical trials for the treatment of diabetic retinopathy, neuropathy, and nephropathy.23 Preliminary evidence demonstrated that RBX may be effective in preventing some of the vascular abnormalities associated with cardiovasculopathies in animal models. In an early study,32 RBX decreased the mitogenic activity of VEGF by selectively inhibiting PKC-b in endothelial cells from bovine aorta. Evidence suggests that the inhibition of PKC by RBX prevents the progression of diabetic nephropathy, including the inhibition of mesangial expansion in a diabetic (db/db) rodent model of type 2 diabetes.33 Although the administration of RBX (10 mg/ kg body weight/day) for 16 weeks to db/db mice had no effect on blood glucose levels, body weight, or kidney weight compared to untreated controls, the treatment did result in inhibition of glomerular PKC-b activation and mesangial expansion and a reduction in urinary albumin excretion rates.33 More recently, RBX treatment (32 mg/day) in adults with type 2 diabetes for 1 year maintained estimated glomerular ltration rat (-2.5 1.9 ml/minute per 1.73 m2; P = 0.185), whereas it was signicantly reduced in the placebo group (-4.8 1.89 ml/minute per 1.73 m2; P = 0.009).34 Because these subjects were already treated with stable doses of an angiotensin-converting enzyme inhibitor and/or angiotensin receptor blocker, the ndings suggested that inhibition of PKC through RBX treatment acts through a distinct pathway offering additional benets to traditional therapies for diabetic nephropathy. In diabetic retinopathy, treatment with RBX inhibited the progression of DME as well as reduced visual loss in clinical studies.35,36 In a 3-year, randomized controlled study35 of 685 subjects with type 1 or type 2 diabetes attending 70 clinics, oral treatment with RBX (32 mg/day) was investigated in patients with moderately severe to very severe non-proliferative retinopathy. Administration of RBX was associated with a 40% reduction in the prevalence of sustained moderate visual loss (sustained from months 30 to 36 of study; 9.1% versus 5.5% in placebo- and RBX-treated subjects, respectively; P = 0.034). There was also a two-fold increase in the baseline-to-endpoint visual improvement of 15 letters (P = 0.005) and inhibition of the progression of clinically signicant macular edema (P = 0.003) compared to placebo control.35 A 30-month follow-up study of subjects with DME demonstrated that daily oral administration of RBX, 32 mg, reduced the pro1532

gression of DME to a sight-threatening stage compared to placebo (P = 0.054).36 ARE PERIODONTAL PATHOLOGIES OF DIABETES MICRO- OR MACROVASCULAR COMPLICATIONS? Diabetic complications are usually separated into micro- or macrovascular diseases because they differ in some of the risk factors. Some of the metabolic abnormalities, such as lipids, fatty acids, and insulin resistance, may be present in obesity and diabetes, whereas hyperglycemia is only present in diabetes. Therefore, the development of the diabetic complications involving cardiovascular or macrovascular diseases are associated with all of the risk factors stated above. However, hyperglycemia is essential for the development of diabetic microvascular diseases but not for cardiovascular complications. This conclusion is derived from the clinical observations that, without hyperglycemia, such as in obese or insulin-resistant people without diabetes, classic lesions of diabetic retinopathy or nephropathy are generally not found. Multiple studies,37-39 including those of Dr. Robert Genco, suggested that the increased risk for periodontal disease exists in patients with obesity without diabetes. These ndings suggested that periodontal disease associated with diabetes may be more related to macrovascular complications of diabetes. This separation could be potentially important for understanding the mechanism of diabetic periodontal disease. Several mechanisms related to diabetic complications could be applicable to micro- and macrovascular diseases of diabetes. These mechanisms include the following: increases in oxidative stress, inammation, AGE formation, and activation of several signaling pathways involving PKC, MAPK, and others. In this issue, the role of inammation and oxidative stress in causing periodontal disease has been reviewed by other authors and, therefore, will not be done here. Increases in oxidative stress and inammatory factors have been clearly identied and shown in subjects with obesity and insulin resistance with or without diabetes. Therefore, it is very likely that similar factors, which cause increases in inammatory molecules and oxidative stress in obese and insulin-resistant states, such as increased fatty acids, could also be causing problems in the periodontal tissue. Furthermore, when diabetes occurs, hyperglycemia would be an additional risk factor that induces oxidative stress and inammation to accelerate dysfunctions in periodontal tissue, which are already present because of obesity or insulin resistance. One example of the specic adverse effects of hyperglycemia, which is not present in insulin-resistant and obese subjects, is its inhibitory effect on neutrophils to resolve infection. Increases in susceptibility

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to infection could accelerate the destruction of periodontal tissues. Clearly, studies are needed to clarify the role of hyperglycemia and other metabolic factors, such as lipids, fatty acids, and the role of insulin resistance, in periodontal tissue metabolism and function. It is very likely that those metabolic changes and insulin actions play an important role in the function and survival of gingiva. In addition, it is also possible that periodontal disease could be a manifestation of an important syndrome, the metabolic syndrome, which manifests multiple metabolic abnormalities that may or may not include diabetes but is strongly associated with insulin resistance. Therefore, it is possible that periodontal disease could be a marker for metabolic syndrome. Reciprocally, it is also possible that people with metabolic syndrome may have an increased risk for the development of periodontal disease. In the screening of patients with metabolic syndrome, data need to be gathered to determine whether the evaluation of periodontal disease should be included. CONCLUSIONS Diabetes and its complications are a growing concern, given the increases in its worldwide prevalence and its association with obesity. Inammatory processes are implicated in the onset of diabetes and the progression of complications. Adiposity, and in particular the release of inammatory cytokines and other mediators from adipose cells, is associated with the development of insulin resistance in type 2 diabetes, partially through activation of the NF-kB pathway. Other factors that activate the NF-kB pathway and are associated with the development of insulin resistance include obesity and a high-fat diet, hyperglycemia, PKC activation, and oxidative stress. Hyperglycemia is the major risk factor for the development of microvascular diabetic complications, such as retinopathy, neuropathy, and nephropathy. Diabetic cardiovasculopathy has a combination of risk factors, including insulin resistance, free fatty acidemia, hypertension, hyperlipidemia, and inammation. Although evidence suggests that periodontal disease is associated with increases in inammatory markers, the major risk factors for its development in diabetes are still unclear. It is important to clarify whether the causes of periodontal disease associated with diabetes are similar to micro- or macrovascular diseases, because the treatment may be different. A well-described pathway underlying the development of diabetic vascular complications is via activation of PKC isoforms, PKC-b in particular. At the level of individual tissues, activation of PKC-b is associated with the development and/or progression of microvascular and macrovascular complications. Administration of the oral agent RBX to selectively inhibit PKC-b activation ameliorated the vascular complications of diabetes. It is a novel

therapy that potentially could be used in conjunction with established agents for diabetic complications. ACKNOWLEDGMENTS This work was supported by grants R01 DK53105 and R01 EY016150 from the National Institutes of Health, Bethesda, Maryland, to GLK, research grant #ADA 1-08-RA-96 from the American Diabetes Association, Alexandria, Virginia, to GLK, and the Diabetes and Endocrinology Research Center at Joslin Diabetes Center, which is funded by National Institutes of Health grant P30 DK36836. The initial draft of this manuscript was developed by a medical writer (Axon Medical Communications Group, Toronto, Ontario) based on content provided solely by the author. The nal manuscript submitted was under the sole control of the author. REFERENCES
1. (CDC) Center for Disease Control and Prevention. Diabetes data & trends. Number (in millions) of persons with diagnosed diabetes, United States, 1980-2005. Available at: http://www.cdc.gov/diabetes/statistics/ prev/national/gpersons.htm. Accessed February 9, 2008. 2. (CDC) Center for Disease Control and Prevention. Diabetes data & trends. Overweight and obesity. U.S. Obesity Trends 1985-2006. Available at: http://www.cdc.gov/ nccdphp/dnpa/obesity/trend/maps/. Accessed February 9, 2008. 3. Wild S, Roglic G, Green A, Sicree R, King H. Global prevalence of diabetes: Estimates for the year 2000 and projections for 2030. Diabetes Care 2004;27: 1047-1053. 4. Soskolne WA, Klinger A. The relationship between periodontal diseases and diabetes: An overview. Ann Periodontol 2001;6:91-98. 5. (CDC) Center for Disease Control and Prevention. National diabetes fact sheet. United States, 2005 General information. Available at: http://apps.nccd.cdc.gov/ DDTSTRS/template/ndfs_2005.pdf. Accessed February 9, 2008. 6. Singh R, Shaw J, Zimmet P. Epidemiology of childhood type 2 diabetes in the developing world. Pediatr Diabetes 2004;5:154-168. 7. Treszl A, Szereday L, Doria A, King GL, Orban T. Elevated C-reactive protein levels do not correspond to autoimmunity in type 1 diabetes. Diabetes Care 2004; 27:2769-2770. 8. Devaraj S, Cheung AT, Jialal I, et al. Evidence of increased inammation and microcirculatory abnormalities in patients with type 1 diabetes and their role in microvascular complications. Diabetes 2007;56: 2790-2796. 9. Vozarova B, Weyer C, Lindsay RS, Pratley RE, Bogardus C, Tataranni PA. High white blood cell count is associated with a worsening of insulin sensitivity and predicts the development of type 2 diabetes. Diabetes 2002;51: 455-461. 10. Pradhan AD, Manson JE, Rifai N, Buring JE, Ridker PM. C-reactive protein, interleukin 6, and risk of developing type 2 diabetes mellitus. JAMA 2001;286: 327-334. 1533

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11. Thorand B, Lowel H, Schneider A, et al. C-reactive protein as a predictor for incident diabetes mellitus among middle-aged men: Results from the MONICA Augsburg cohort study, 1984-1998. Arch Intern Med 2003;163:93-99. 12. Mooradian AD. Obesity: A rational target for managing diabetes mellitus. Growth Horm IGF Res 2001; 11(Suppl. A):S79-S83. 13. Shoelson SE, Lee J, Goldne AB. Inammation and insulin resistance. J Clin Invest 2006;116:17931801. 14. Hotamisligil GS. Inammation and metabolic disorders. Nature 2006;444:860-867. 15. Petersen KF, Dufour S, Befroy D, Garcia R, Shulman GI. Impaired mitochondrial activity in the insulinresistant offspring of patients with type 2 diabetes. N Engl J Med 2004;350:664-671. 16. Albert MA, Ridker PM. C-reactive protein as a risk predictor: Do race/ethnicity and gender make a difference? Circulation 2006;114:e67-e74. 17. Hotamisligil GS, Shargill NS, Spiegelman BM. Adipose expression of tumor necrosis factor-alpha: Direct role in obesity-linked insulin resistance. Science 1993;259: 87-91. 18. Ofei F, Hurel S, Newkirk J, Sopwith M, Taylor R. Effects of an engineered human anti-TNF-alpha antibody (CDP571) on insulin sensitivity and glycemic control in patients with NIDDM. Diabetes 1996;45: 881-885. 19. Yuan M, Konstantopoulos N, Lee J, et al. Reversal of obesity- and diet-induced insulin resistance with salicylates or targeted disruption of Ikkbeta. Science 2001;293:1673-1677. 20. Fleischman A, Shoelson SE, Bernier R, Goldne AB. Salsalate improves glycemia and inammatory parameters in obese young adults. Diabetes Care 2008; 31:289-294. 21. Tsai C, Hayes C, Taylor GW. Glycemic control of type 2 diabetes and severe periodontal disease in the US adult population. Community Dent Oral Epidemiol 2002;30:182-192. 22. Scott JA, King GL. Oxidative stress and antioxidant treatment in diabetes. Ann N Y Acad Sci 2004;1031: 204-213. 23. He Z, King GL. Protein kinase C beta isoform inhibitors: A new treatment for diabetic cardiovascular diseases. Circulation 2004;110:7-9. 24. Dalla Vestra M, Mussap M, Gallina P, et al. Acutephase markers of inammation and glomerular structure in patients with type 2 diabetes. J Am Soc Nephrol 2005;16(Suppl. 1):S78-S82. 25. Izuora KE, Chase HP, Jackson WE, et al. Inammatory markers and diabetic retinopathy in type 1 diabetes. Diabetes Care 2005;28:714-715. 26. Meleth AD, Agron E, Chan CC, et al. Serum inammatory markers in diabetic retinopathy. Invest Ophthalmol Vis Sci 2005;46:4295-4301.

27. Das Evcimen N, King GL. The role of protein kinase C activation and the vascular complications of diabetes. Pharmacol Res 2007;55:498-510. 28. He Z, King GL. Can protein kinase C beta-selective inhibitor, ruboxistaurin, stop vascular complications in diabetic patients? Diabetes Care 2005;28:2803-2805. 29. Ohshiro Y, Ma RC, Yasuda Y, et al. Reduction of diabetes-induced oxidative stress, brotic cytokine expression, and renal dysfunction in protein kinase C beta-null mice. Diabetes 2006;55:3112-3120. 30. Aiello LP, Avery RL, Arrigg PG, et al. Vascular endothelial growth factor in ocular uid of patients with diabetic retinopathy and other retinal disorders. N Engl J Med 1994;331:1480-1487. 31. Jirousek MR, Gillig JR, Gonzalez CM, et al. (S)-13[(dimethylamino)methyl]-10,11,14,15-tetrahydro-4, 9:16, 21-dimetheno-1H, 13H-dibenzo[e,k]pyrrolo[3, 4-h][1,4,13]oxadiazacyclohexadecene-1,3(2H)-d ione (LY333531) and related analogues: Isozyme selective inhibitors of protein kinase C beta. J Med Chem 1996; 39:2664-2671. 32. Xia P, Aiello LP, Ishii H, et al. Characterization of vascular endothelial growth factors effect on the activation of protein kinase C, its isoforms, and endothelial cell growth. J Clin Invest 1996;98:2018-2026. 33. Koya D, Haneda M, Nakagawa H, et al. Amelioration of accelerated diabetic mesangial expansion by treatment with a PKC beta inhibitor in diabetic db/db mice, a rodent model for type 2 diabetes. FASEB J 2000; 14:439-447. 34. Tuttle KR, Bakris GL, Toto RD, McGill JB, Hu K, Anderson PW. The effect of ruboxistaurin on nephropathy in type 2 diabetes. Diabetes Care 2005;28:26862690. 35. Aiello LP, Davis MD, Girach A, et al. Effect of ruboxistaurin on visual loss in patients with diabetic retinopathy. Ophthalmology 2006;113:2221-2230. 36. PKC-DMES Study Group. Effect of ruboxistaurin in patients with diabetic macular edema: Thirty-month results of the randomized PKC-DMES clinical trial. Arch Ophthalmol 2007;125:318-324. 37. Al-Zahrani MS, Bissada NF, Borawski EA. Obesity and periodontal disease in young, middle-aged, and older adults. Journal of Periodontology 2003;74:610-615. 38. Genco R, Grossi S, Ho A, Nishimura F, Murayama Y. A proposed model linking inammation to obesity, diabetes, and periodontal infections. Journal of Periodontology 2005;76:2075-2084. 39. Ritchie C. Obesity and periodontal disease. Periodontology 2000 2007;44:154-163. Correspondence: Dr. George L. King, Joslin Diabetes Center and Joslin Clinic, One Joslin Place, Boston, MA 02215. Fax: 617/732-2637; e-mail: george.king@joslin.harvard.edu. Submitted May 5, 2008; accepted for publication May 30, 2008.

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